A new study, entitled “Genome-wide methylation profiling of the different stages of hepatitis B virus-related hepatocellular carcinoma development in plasma cell-free DNA reveals potential biomarkers for early detection and high-risk monitoring of hepatocellular carcinoma” may lead to next-generation diagnostic models for the disease. The study was published in Clinical Epigenetics by Yangxing Zhao and Feng Xue, first co-authors of the study, and led by Dr. Jian Yu from the Shanghai Cancer Institute, and Dr. Qiang Xia from Shanghai Jiao Tong University.

In Southeast Asia, a relevant model of hepatocellular carcinoma (HCC) has been described that includes all the stages of the disease from chronic hepatitis B infection (CHB) to liver cirrhosis (LC) and, ultimately, to HCC. Most tumors when developing acquire specific biological phenotypes; epigenetic modifications, like DNA methylation, that seem to play crucial roles for the development of the tumor. A genome-wide methylation profiling of plasma cell-free DNA (cfDNA) has never been done to investigate the development of HCC.

The research team used MethylCap-seq, a profiling method that enables the identification of the methylated portion of the genome and reveals new regions that are differently methylated in a biological sample. The authors analyzed the genome-wide plasma cell-free DNA methylation profiles from the pool of samples of healthy control (HC), chronic hepatitis B infection (CHB), liver cirrhosis (LC) and hepatocellular carcinoma (HCC) patients. Then they independently validated the library data for the DNA of the tissue and plasma cell-free DNA using three methods: 1) methylation-specific PCR (MSP), the most common technique to study CpG island methylation, 2) quantitative Methylation-Specific PCR (qMSP), a method designed to amplify bisulphite-converted methylated DNA target sequences in the presence of an excess of unmethylated sequences, 3) and Multiplex-BSP-seq.

In this study, the authors found that dynamic characteristics of plasma cell-free DNA methylation matched the normal development of HCC. The results identified the presence of 240, 272 and 286 genes differentially methylated (DMGs) related, respectively, to the early, middle and late stages of progression of hepatocellular carcinoma. In addition, three differentially methylated genes were identified, ZNF300, SLC22A20 and SHISA7, that possibly enable to distinguish between chronic hepatitis B infection and liver cirrhosis and between liver cirrhosis and hepatocellular carcinoma.

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